Relative Risk, One Result at a Time

When research suggests that a single chemical may cause harm, public concern rises, as it has for the plastic additive bisphenol A (BPA) in recent years. But many more of the 83,000 or so humanmade chemicals used in the United States receive little attention. The possible effects of chemicals in combination get still less scrutiny, even though the potential that some chemicals will interact is high, given their numbers.

This may be due in part to the staggering amount of work required to discern those effects. It would be a very difficult task to keep up with research on all of these substances, much less evaluate their relative risk as new results appear. The U.S. Environmental Protection Agency (EPA) has put considerable effort into this under the Toxic Substances Control Act, but the Act has not been updated since its passage in 1976 and excludes many substances from the agency’s purview.

Substances that have the potential to disrupt development in an organism are of special concern. The results of exposure to such chemicals can range from birth defects to developmental irregularities that don’t appear until later in life. Determining whether a substance is an endocrine disruptor, how strongly it acts and at what concentrations, not to mention deciphering hormone pathways themselves, takes a great deal of time and resources. Studies in the lab can’t be directly extrapolated to real-life situations, but they can offer clues about new routes to explore, along with help in evaluating the risk posed by various chemicals.

Heather Patisaul, a biologist at North Carolina State University, studies the effects of BPA and other compounds suspected to disrupt hormonal processes, using female rats as models. “The biggest unknown,” she says, “is if human harm is indeed resulting from exposure to these chemicals at low doses. If it is, it requires a major paradigm shift in how we approach toxicology, because the current strategies are ill equipped to deal with endocrine disruptors.”

A new study adds several more pieces to the puzzle. In a September 2011 study in the Proceedings of the National Academy of Sciences of the U.S.A., Eunah Chung, Maria C. Genco, Laura Megrelis and Joan V. Ruderman chose a less known, but widely used, substance to investigate: triclocarban (3,4,4’-trichlorocarbanilide, or TCC).

TCC has been used as an antimicrobial in consumer products since the 1950s. A 2001 study found that it was present in 84 percent of antimicrobial bar soaps sold in the United States. It’s often mentioned in the same breath with triclosan: Both are halogenated carbons used in soaps and other products, but their chemical identities are unique. The EPA reports that between 1 and 10 million pounds of TCC were used in the United States in 2002. People who shower with soap containing TCC absorb it through their skin. It is metabolized quickly by humans but persists in surface waters and in sewage sludge that is spread on agricultural fields.

Ruderman and her coauthors looked at the gene aromatase-B (AroB) in the brains of developing zebrafish embryos. AroB is regulated by estrogen, among other compounds, and is expressed in subregions of the brain including the hypothalamus and preoptic areas. To determine what concentration of TCC to use, they tested a range, then chose one that did not show signs of developmental delay or toxicity. The 0.25 micromolar experimental concentration was about 1600 times higher than a high-end estimate of levels in surface waters in an industry report to the EPA, and about 12 times higher than a high-end estimate from a university-based study in Environmental Health Perspectives.

The team found that TCC had little effect on AroB when introduced without estrogen, but that it strongly enhanced the effects of introduced estrogen on the gene, with a twofold greater increase than that induced by estrogen alone. They also tested the effects of BPA and found that it induced the gene’s expression even without estrogen present.

Then the researchers exposed embryos to TCC and BPA together. Rather than amplifying the effect of the estrogen-mimicking BPA, TCC suppressed it: Its presence along with BPA resulted in about a twofold decrease in transcription of aromatase-B compared to embryos exposed to BPA alone.

“The experiments we did with BPA plus TCC were an example where each one has a positive effect on an estrogenlike process,” Ruderman says. “But you put them together and they are not additive—in fact in some ways they suppress each other.” It’s surprising that TCC would amplify estrogen’s effect but reduce the effect of an estrogen mimic—a reminder that chemicals in combination can act unpredictably.

In a 2008 study in Environmental Health Perspectives, Bruce Hammock, an entomologist at the University of California, Davis, and colleagues found that TCC enhanced estrogen- and testosterone-dependent gene expression by 2.5 times in human cells. “The major significance” of Ruderman’s study, he says, “is an elegant demonstration that there is the potential for two known environmental chemicals to synergize for an enhanced biological effect.” He thinks it’s unlikely that environmental exposure to both chemicals will be high enough to create such effects, but he notes, “This is a cautionary tale in terms of mixtures in general. As a society we are using thousands of high-volume chemicals with little regard to environmental or human health effects.”

Andreas Kortenkamp, a mixtures toxicologist at Brunel University, thinks that scientists can help interpret incomplete or inconclusive evidence so that policy makers can move forward: “Because of these many unknowns and uncertainties, we have to develop ways of making very important decisions on the basis of incomplete knowledge. That’s not easy, and we as scientists can actually make a little contribution there.”

Such contributions are especially important because the EPA relies in part on voluntary industry reporting to glean information. It has asked companies to make information about high-production-volume (HPV) chemicals available, under its HPV Challenge program, in order to determine their relative priority. A 2009 HPV Challenge assessment, submitted by the Triclocarban Consortium (whose members include the Bayer Corporation and Clariant Corporation), stated that “there is negligible likelihood of harm to man and the environment” from the manufacture and use of TCC, calling it “a low priority for further work.” In the EPA’s subsequent initial risk-based prioritization for TCC, based on the Consortium’s assessment along with other data, it listed the substance as high priority, noting its high potential risk to aquatic organisms from environmental releases—a risk documented in the Consortium assessment. That risk is also foremost in Ruderman’s and Hammock’s minds. “You’ve got to wonder if TCC is affecting estrogen signaling in aquatic organisms out there,” Ruderman says.

The EPA and TCC Consortium reports marked the potential risk to the general population as low. But Ruderman asks, “What is the molecular mechanism by which TCC is acting to affect estrogen signaling? Clearly it’s having an effect, and if it’s affecting estrogen signaling, might it not affect other hormones or other cell systems in bad ways that we don’t know about?”

Hammock is less worried about TCC than he is about several flame-retardant chemicals used to meet the standards of a California law regulating furniture and bedding, and U.S. flame-resistance standards for computer circuit boards. “Chemicals aren’t fundamentally bad or good,” he says, “but they do have properties, and in general, halogens on carbon molecules in high-volume chemicals usually have come back to bite us. Sometimes it’s fifteen years later, but we usually run into trouble. Fire retardants fall into that class of compounds, and some personal-care products too.”

The potential combinatory effects of such chemicals have been so little studied that more surprises are sure to come. It would be good, Ruderman says, to adhere to the precautionary principle embraced by the field of public health. The best hope she sees for that is informing women who are pregnant or who plan to have children: “People will stop all sorts of unhealthy behaviors for the health of their unborn child.” She recommends the site http://safecosmetics.org as a resource.

“It sounds terrific, right?” she says. “Why wouldn’t you want something that’s antibacterial? But it turns out that plain soap is just as good in getting rid of germs. And why would you want to have this other chemical in there if it’s not needed?”